The long-term goal of this project is the characterization of neurotransmitter receptor-mediated information transduction, and its regulation, across neuronal membranes. The primary, but not exclusive, model systems under investigation are those for dopamine receptors. In order to characterize dopamine receptors at the biochemical and molecular levels and study their regulation, there are two major interrelated lines of research which are ongoing: 1 ).investigation of the cell biology, function and regulation of the receptors at the protein level; and 2) the molecular cloning of the receptor genes and investigation of gene structure and regulation in normal and pathophysiological states. 1. Cell Biology and Regulation of Dopamine Receptors. Characterization of the functional and regulatory properties of Dl and D2 dopamine receptors on various neuroblastoma, retinoblastoma, and cDNA-transfected cell lines were continued. The Dl receptors were shown to undergo a cAMP-mediated form of desensitization involving both functional uncoupling and down regulation of the receptors. D2 receptors were also shown to undergo desensitization and down-regulation in response to agonist treatment. A variety of fluorescent, affinity, and antibody probes for dopamine receptors were developed. The fluorescent probes and antibodies were used to map the expression of the Dl, D2 and D3 receptor proteins at the cellular level in the CNS. 2. Molecular Cloning of Dopamine and Other Receptors. A cDNA encoding the DlA dopamine receptor linked to the stimulation of adenylyl cyclase in rat striatum was cloned and expressed in CHO cells. A second Dl B receptor linked to adenylyl cyclase in rat kidney was also identified and cloned. Work continued on the cloning of a third "Dl like" receptor which apparently is linked to the stimulation of phosphatidyl inositol turnover and mobilization of calcium. The genes for both the DlA, D1B, and D2 receptors were isolated. The rat brain Al adenosine receptor was also cloned and expressed. Several other cDNA clones encoding a putative serotonin receptor as well as several "orphan" G protein linked neurotransmitter receptors were identified.